Current coronavirus (SARS-CoV-2) epidemiological, diagnostic and therapeutic approaches: An updated review until June 2020

Coronaviruses are a group of enveloped viruses with non-segmented, single-stranded, and positive-sense RNA genomes. In December 2019, an outbreak of coronavirus disease 2019 (COVID-19) caused by the novel severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), in Wuhan City, China. The World Health Organization (WHO) declared the coronavirus outbreak as a global pandemic in March 2020. Fever, dry cough and fatigue are found in the vast majority of all COVID-19 cases. Early diagnosis, treatment and future prevention are keys to COVID-19 management. Currently, the unmet need to develop cost-effective point-of-contact test kits and efficient laboratory techniques for confirmation of COVID-19 infection has powered a new frontier of diagnostic innovation. No proven effective therapies or vaccines for SARS-CoV-2 currently exist. The rapidly increasing research regarding COVID-19 virology provides a significant number of potential drug targets. Remdesivir may be the most promising therapy up till now. On May 1, 2020, Gilead Sciences, announced that the U.S. Food and Drug Administration (FDA) has granted emergency use authorization (EUA) for the investigational Remdesivir as a potential antiviral for COVID-19 treatment. On May 7, 2020, Gilead Sciences, announced that the Japanese Ministry of Health, Labour and Welfare (MHLW) has granted regulatory approval of Veklury® (Remdesivir) as a treatment for SARS-CoV-2 infection, the virus that causes COVID-19 acute respiratory syndrome, under an exceptional approval pathway. Also, Corticosteroids are recommended for severe cases only to suppress the immune response and reduce symptoms, but not for mild and moderate patients where they are associated with a high-risk side effect. Based on the currently published evidence, we tried to highlight different diagnostic approaches, side effects and therapeutic agents that could help physicians in the frontlines.


INTRODUCTION
In December 2019, a novel coronavirus, SARS-CoV-2, was identified as the pathogen causing coronavirus disease  in Wuhan, China. On March 11, 2020, the World Health Organization declared COVID-19 as a global pandemic (Whitworth, 2020).
COVID-19 is an enveloped, positivesense, single-stranded RNA virus that belongs to the beta-CoV genus, which also includes SARS-CoV and MERS-CoV. It shares 89 % nucleotide identity with bat SARS-like CoVZXC21 and 82 % identity with human SARS-CoV (Chan et al., 2020a).
COVID-19 is transmitted by inhalation or contact with infected droplets. The incubation period for COVID-19 is on average, 5-6 days, but can be up to 14 days. During this period, also known as the "presymptomatic" period, some infected persons can be contagious, from 1-3 days before symptom onset (Wei et al., 2020). The clinical manifestations of COVID-19 varied from asymptomatic carrier status, acute respiratory disease (ARD) and pneumonia. The prevalence of asymptomatic cases is significant (20-86 % of all infections) and is defined as individuals with positive viral nucleic acid tests but without any COVID-19 symptoms. Most people with COVID-19 develop only mild (40 %) or moderate (40 %) disease, approximately 15 % develop a severe disease that requires hospitalization and oxygen support, and 5 % have a critical disease with complications such as respiratory failure, acute respiratory distress syndrome (ARDS), sepsis and septic shock, thromboembolism, and/or multiorgan failure, including acute kidney injury and cardiac injury (CDC, 2020b) Older age, co-morbidities such as diabetes, hypertension, cardiac disease, chronic lung disease, cancer and BMI > 40 kg/m 2 have been reported as risk factors for severe disease and death (CDC, 2020a). Wang and colleagues (2020a) reported that there are 6 common signs and symptoms that 30 % of the patients have felt including fever (98.5 %), fatigue (69.9 %), dry cough (59.4 %), anorexia (39.8 %), myalgia (34.8 %), dyspnea (31.1 %) and for the most common comorbidities are hypertension (31.1 %) and cardiovascular disease (14.5 %). Symptoms may develop 2 days to 2 weeks following exposure to the virus (CDC, 2020b). According to Wu and McGoogan (2020), among 72,314 SARS-CoV-2 cases reported to the Chinese Center for Disease Control and Prevention (CCDC), 81 % were mild (mild or absent pneumonia), 14 % were severe (dyspnea, hypoxia, > 50 % lung involvement within 1-2 days), 5 % were critical (respiratory failure, shock, multiorgan dysfunction), and 2.3 % were fatal. Symptoms in children with infection appear to be uncommon, although some children with severe COVID-19 have been reported (CDC, 2020a). Based on currently available information and clinical expertise, risk factors for severe COVID-19 include older adults ≥ 65 years as well as people of all ages with chronic lung disease or moderate to severe asthma, serious heart conditions, diabetes, severe obesity, chronic kidney disease, liver disease and immunocompromised people (CDC, 2020a).

SUGGESTED INFECTION MECHANISM
Upon infection with COVID-19, it binds to the host cell's angiotensin-converting enzyme 2 (ACE2) receptors which commonly expressed on the epithelial cells of alveoli, trachea, bronchi, and bronchial serous glands of the respiratory tract. Then the virus enters and replicates in these cells (Liu et al., 2011). The newly developed virions are then released and infect new target cells. Unfortunately, there is no specific antiviral treatment or vaccine recommended for COVID-19 that is currently available.

SARS-COV-2 DIAGNOSIS
The diagnosis of COVID-19 mainly depends on the demonstration of the virus in respiratory secretions by special molecular tests.
Common laboratory findings include normal/ low white cell counts with elevated C-reactive protein (CRP). The computerized tomographic chest scan is usually abnormal even in those with no symptoms or mild disease (Singhal, 2020). In addition to laboratory testing capacity and reagent shortages, the rapidly growing SARS CoV 2 pandemic has encouraged many diagnostic manufacturers to develop and sell fast and easy-to-use equipment to facilitate testing outside the laboratory (WHO, 2020a). Currently, there are two main categories commercially available for COVID-19 tests. The first category includes molecular assays for detection of SARS-CoV-2 viral RNA using polymerase chain reaction (PCR)-based methods. The second category includes serological and immunological assays that largely depend on detecting antibodies produced by individuals as a result of exposure to the virus or on the detection of antigenic proteins in infected individuals. It is necessary to ensure that these two categories of tests serve overlapping purposes in the management of the SARS-CoV-2 pandemic (Carter et al., 2020). Current COVID-19 diagnostic tools and techniques are shown in Table 1 and a diagnostic model for COVID-19 in Figure 5.

SARS-COV-2 DIFFERENT THERAPEUTIC APPROACHES
Symptomatic treatment and oxygen therapies represent the major treatment interventions for patients with severe infection. Mechanical ventilation may be necessary in cases of respiratory failure refractory to oxygen therapy, whereas hemodynamic support is essential for managing septic shock (Cascella et al., 2020).
To the best of our knowledge, different therapeutic approaches have been evaluated against COVID-19 in vivo, vitro and in clinical trials. Many of these therapies had a great impact on clinical recovery. Current COVID-19 therapies are shown in Table 2. Both SARS and SARS-CoV-2 invade the cell through the ACE2 receptor. SARS-CoV reduces ACE2 expression, causing an imbalance between the ACE/Ang II/AT1R axis and the ACE2/Ang (1-7)/Mas receptor axis. A novel therapeutic strategy for hypertension targets the ACE/Ang II/AT1R axis. Angiotensin-Converting Enzyme Inhibitors (ACEIs) and agents acting on the renin-angiotensin system (ARAS) inhibit the ACE/Ang II/AT1R pathway in addition to modulation of the ACE2/Ang (1-7)/Mas receptor pathway. COVID-19 patients observed to have a dysfunction in the renin-angiotensin system (RAS). It was also noticed that ACEI/angiotensin-receptor blockers (ARB) had the potential to decrease the viral load, regulate immune function and inhibit inflammatory responses. MSCs transplantation improved the outcome of COVID-19 patients. These findings may be due to regulating inflammatory response and promoting tissue repair and regeneration, where mesenchymal stem cells are blank cells that can differentiate into most cellular types in addition to their paracrine fashion of cytokines and growth factors that dampen inflammation and cell death. Leng et al., 2020 9 Tocilizumab Well-known recombinant humanized anti-human interleukin-6 receptor monoclonal antibody that is mainly used for rheumatoid arthritis patients. In COVID-19 infection, a massive number of T-lymphocytes and mononuclear macrophages are activated, emitting different cytokines such as interleukin-6 (IL-6), which binds to the IL-6 receptor on its target cells, causing the cytokine storm and severe inflammatory responses in most organs including lungs, liver, kidney and other tissues and organs. Tocilizumab can specifically bind soluble interleukin-6 receptor (sIL-6R) and membrane-bound interleukin-6 receptor (mIL-6R) and inhibits their signal transduction.

Xu et al., 2020a
10 Human monoclonal antibody Coronavirus neutralizing antibodies target the viral trimeric spike (S) glycoproteins that exist on the viral surface and mediate viral entry into the host cells. Li et al., 2020 11 Baricitinib Most viruses invade cells through receptor-mediated endocytosis. ACE2 is the receptor that COVID-19 uses to infect lung cells. ACE2 is a cell surface protein that distributed many cells including kidney, blood vessels, heart, and, especially, lung AT2 alveolar epithelial cells. AT2 cells are mainly prone to viral infection. One of the known regulators of endocytosis is the AP2-associated protein kinase 1 (AAK1). Disruption of AAK1 might, in turn, interrupt the passage of the virus into cells and also the intracellular assembly of virus particles. Baricitinib with therapeutic dosage (2 mg or 4 mg once daily) is sufficient to inhibit AAK1.

Richardson et al., 2020
12 COVID-19 recovered patients' convalescent plasma COVID-19 recovered patients' convalescent plasma contains a huge quantity of COVID-19 monoclonal antibodies. So, direct administration of COVID-19 recovered patients' convalescent plasma might suppress viremia. Several studies showed a shorter hospital stay and lower mortality rate in convalescent plasma-treated patients than those who were not treated with it.
Chen et al., 2020 Nelfinavir was recommended to be used as a potential therapy for COVID-19 through inhibition of its main protease using an integrative approach combining molecular docking, homology modeling and binding free energy calculation.  15 Regulation of interferon production The DNA sensor cyclic GMP-AMP synthase (cGAS), anaplastic lymphoma kinase (ALK) and stimulator of interferon genes (STING) were suggested to be potential therapeutic effective targets preventing the cytokine storm during COVID-19 infection. HUCMSCs have shown significant tissue repair and immunomodulation with a low immunogenic effect that makes these cells very ideal candidates for the allogenic adoptive transfer therapy. HUCMSCs were also suggested to be a potential treatment for H5N1 infectioninduced acute lung injury. COVID-19 showed a similar inflammatory cytokine profile to that of H5N1. Liang et al., 2020 18 CD-sACE2 Inclusion Compounds The main receptors for SARS-CoV and SARS-CoV-2 are ACE2 Soluble ACE2 (sACE2) retaining ACE2 enzyme activity in addition to binding SARS-CoV S-protein. So sACE2 can inhibit SARS-CoV infected cells. Since SARS-CoV and SARS-CoV-2 infection mechanisms are the same, sACE2 can inhibit the infection of SARS-CoV-2.
To improve the water solubility of sACE2, the formation of a complex between CD and sACE2 would be effective and enables it to meet drug atomization inhalation requirements. Sun et al., 2020b 19 Favipiravir (Avigan®) Favipiravir is a Pyrazine carboxamide broad-spectrum antiviral drug that has been approved in Japan for influenza treatment. It is a prodrug that is phosphorylated and ribosylated intracellularly to form its active metabolite (Favipiravir ibofuranosyl -5′-triphosphate) that acts as a competitive inhibitor for viral purine nucleosides in addition to inhibition of RNA-dependent RNA polymerase (RdRp) of RNA viruses. Finally, it interferes with viral replication. Du and Chen, 2020 20 Ivermectin FDA-approved for parasitic infections treatment. Caly et al. reported that Ivermectin has the potential to inhibit COVID-19 in vitro by interfering with the nuclear import of host and viral proteins, where its single treatment was able to cause ~5000-fold reduction in COVID-19 virus after 48 h in cell culture model. Orally bioavailable prodrug (β-d-N 4 -hydroxycytidine-5'-isopropyl ester) has been reported to improve pulmonary function and reduces COVID-19 titer through induction of transition mutation frequency in viral RNA causing lethal mutagenesis of COVID-19. Sheahan et al., 2020 22 Atazanavir Atazanavir inhibits the activity of COVID-19 essential protease, causing a decline of viral replication in addition to its ability to stop the cytokine storm-associated mediator releasing. So, it acts both as an anti-inflammatory and antiviral candidate.

Recombinant Human Erythropoietin
It has a protective effect on lung tissue by inhibiting NF-κB expression in lung tissues, inhibition of IL-6 and TNF-alpha as proinflammatory cytokines and induction of anti-inflammatory cytokine IL-10. Hadadi et al., 2020 24 Ribavirin It is a guanosine analog that interferes with RNA and DNA virus's replication through interference with viral polymerases and interference with RNA capping.
It was shown to have limited value for the treatment of COVID-19 as monotherapy with high cytotoxicity but when used in combination with other agents it provided the best chance for clinical efficacy. Graci and Cameron, 2006;Sanders et al., 2020 25 Corticosteroids Corticosteroids are mainly used for decreasing the host inflammatory responses into the lungs which may lead to acute lung injury and acute respiratory distress syndrome (ARDS) but corticosteroids may have adverse effects, including delayed viral clearance and the high risk of secondary infection. Moreover, direct evidence for corticosteroids in the treatment of COVID-19 is limited. Russell et al., 2020 26 Lianhuaqingwen (Anti-Viral and Anti-Inflammatory) Traditional Chinese medicine that has been previously used for influenza treatment with broad-spectrum anti-influenza effects. Lianhuaqingwen was found to inhibit COVID-19 replication in vitro with a significant reduction in pro-inflammatory cytokines (TNF-α, IL-6, CCL-2/MCP-1 and CXCL-10/IP-10).

Runfeng et al., 2020
27 Anticoagulant treatment Low molecular weight heparin appears to be associated with better prognosis in severe COVID-19 patients meeting SIC criteria or with markedly elevated D-dimer. An interventional clinical trial on 128 participants in Xijing Hospital showed that the early use of aspirin in COVID-19 patients, which has the effects of inhibiting virus replication, anti-platelet aggregation, anti-inflammatory and anti-lung injury, is expected to reduce the incidence of severe and critical patients, shortens the length of hospital duration and reduces the incidence of cardiovascular complications. Tang et al., 2020; clinicaltrials.gov, 2020

SARS-COV-2 THERAPEUTIC APPROACHES -SIDE EFFECTS
Despite the approved beneficial effects of these therapeutic approaches, recent studies concluded that most of these candidate's administration has a toxic effect in overdoses, causing common and severe adverse effects including nausea, pruritus, arrhythmias, hypoglycemia, anemia, jaundice, hyper-lipidemia, electrolyte abnormalities, acute renal injury, hematological disorders, hyperuricemia, neuropsychiatric effects and various drug-drug interactions.
Lopinavir/Ritonavir (LPV/r) combination has been reported to have gastrointestinal disorders, so in some SARS-CoV-2 patients, the treatment was stopped due to these severe side events (Owa and Owa, 2020). Notwithstanding the minimal side effects of Teicoplanin, it may cause thrombocytopenia in some treated cases (Terol et al., 1993).
An exploratory randomized controlled trial assessing the efficacy and safety of Arbidol in COVID-19 patients reported that patients had adverse events including diarrhea, nausea and loss of appetite (Eikenberry et al., 2020), also hypotension, acute renal injury, teratogenicity, hypersensitivity, electrolyte abnormalities, fatigue, diarrhea, weakness, anemia and chest pain are the most common risk factors during treatment of COVID-19 patients using inhibitors of the renin-angiotensin system (Ingraham et al., 2020). Zhang and colleagues (2020) reported that intravenous transplantation of Wharton's jelly derived mesenchymal stem cells (hWJCs) was safe and effective especially, in COVID-19 critical severe cases. Regarding Tocilizumab that was used as a treatment for severe COVID-19 cases, it may cause serious adverse reactions, like intestinal perforation, candidiasis and lipid metabolism abnormalities (Tao et al., 2020).
FDA has approved convalescent plasma therapy in COVID-19 critical patients, but up till now, only three studies with small sample size reported effectiveness and safety so more clinical trials are needed to ensure both safety and efficacy (Bloch et al., 2020).
Otherwise Direct-acting antivirals (DAAs) demonstrated, a safe therapeutic approach with common side effects including fatigue, headache, nausea and neuropsychiatric symptoms (Medeiros et al., 2017). Concerning using of Favipiravir (Avigan®) as a treatment for COVID-19 patients, it was reported that Favipiravir elevates plasma uric acid, so this finding should be considered in hyperuricemia, gout and kidney impairment patients (Mishima et al., 2020).
Despite the beneficial effect of Corticosteroids with COVID-19 patients, they are associated with a high risk of death, side effects like bacterial infections and hypokalemia so they are not recommended for mild and moderate COVID-19 patients, but they should be used in severe cases only to suppress the immune response and reduce symptoms .

CHLOROQUINE TRIGGERS OXIDATION AND HEMOLYTIC ANEMIA IN G6PD DEFICIENT CASES & WORLD HEALTH ORGANIZATION DISCONTINUED ITS TREATMENT TRIALS
Glucose-6-phosphate dehydrogenase (G6PD) deficiency is one of the most common human enzymatic disorders affecting around 400 million people worldwide (Luzzatto and Arese, 2018). Decreased G6PD production results in low levels of NADPH and reduced glutathione stimulating hemolytic anemia which is characterized by oxidative stress and red blood cell lysis (Francis et al., 2013).
The risk of hemolytic anemia should be considered during Chloroquine/Hydroxy Chloroquine (CQ/HCQ) therapy of patients with G6PD deficiency (Mohammad et al., 2018). Beauverd and colleagues (2020) reported that SARS-CoV-2 infection can enhance severe acute hemolysis in patients with G6PDdeficiency, and CQ/HCQ can worsen this crisis. During the treatment of SARS-CoV-2, it is important to carefully monitor potential hemolytic effects of CQ/HCQ in G6PD deficiency cases. If a decline in hemoglobin levels during the first days of CQ/HCQ treatment is observed, the treatment should be stopped. Hemolysis usually is reversible after finishing therapy with CQ/HCQ (De Franceschi et al., 2020). Also, Kapoor and Kapoor (2020) warned of the use of CQ because of the risk of hematological disorders in patients with G6PD deficiency.
In contrast, both (Youngster et al. 2010;Beutler 1994) concluded that CQ or HCQ mono-therapies are safe also in G6PD deficient cases. Afra and colleagues (2020) reported that infections might be the most common causes of hemolysis in G6PD deficient patients. Thus, SARS-CoV-2 patients may show significant hemolysis even before CQ or HCQ administration.
Finally, SARS-CoV-2 treatment using CQ or HCQ, especially in areas with high G6PD deficiency prevalence, should alert medical staff to this possible harmful effect. The US Food and Drug Administration warned of cardiotoxicity caused by hydroxychloroquine and mentioned G6PD as a baseline test before the onset of hydroxychloroquine treatment (FDA, 2020). Moreover, in July 2020 the WHO discontinued clinical trials with hydroxychloroquine and lopinavir/ritonavir treatment arms for COVID-19 (WHO, 2020b), where both therapies produced little and no reduction in the mortality of hospitalized SARS-CoV-2 cases when compared to standard of care.

CONCLUSION
Finally, COVID-19 pandemic is a highly infectious disease caused by the novel coronavirus SARS-CoV-2 that can be transmitted through droplets and close contact and repre-sents a global public health crisis. Fever, fatigue and dry coughs are the most common signs and symptoms of COVID-19. Due to rapid transmission, countries around the world should increase attention to disease surveillance systems. SPR gold nanoparticlebased biosensors may be a promising diagnostic technique as it had high sensitivity, selectivity, reliability, portability, is rapid and cheap, but this method is an indirect method, where it detects antibody, so developing of SPR biosensor to detect COVID-19 itself still is a great challenge. No proven effective therapies or vaccines for SARS-CoV-2 currently exist. The most promising therapy up till now maybe Remdesivir, also we recommend Corticosteroids therapy for severe cases only to suppress the immune response and reduce symptoms, but not for mild and moderate patients where they are associated with high-risk side effects. G6PD should be considered as a baseline test for starting CQ or HCQ treatment protocol to avoid its possible hemolytic effect. We should further strive to develop specific medications, support the research and development of vaccines, and also decrease morbidity and death of SARS-CoV-2 to preserve the population.

Authors contribution
Ahmed